While highly active antiretroviral therapy (HAART) is helping many people with HIV/AIDS live longer, healthier lives, these combination or “cocktail” treatments are not always effective and often cause serious adverse side effects. Even among those who do well on HAART, many patients experience treatment failure within a year or two, often because HIV has developed resistance to the drugs used to inhibit its growth. Drug resistant HIV tends to progressively develop resistance to other drugs in the same class, or even to the entire class of drugs. In addition, many people newly infected with HIV may carry viral strains already resistant to current treatments.
Confounding the resistance challenge is the latency of HIV infection. Despite the success of HAART, medical reports suggest that antiretroviral therapy alone is unable to eliminate the viral infection because the virus can persist in a latent form when infected CD4+ lymphoblasts carrying an integrated copy of the HIV-1 genome revert back to a resting memory state. In this state, CD4 cells are minimally permissive for virus gene expression, and infected memory cells can survive for many years. Following reexposure to the relevant antigen or other activating stimuli, these cells can begin to produce virus again. Though HAART may be effective for an acute infection, it appears unable to eliminate this viral reservoir, which can serve as a permanent archive for wild-type virus and for drug-resistant variants that arise during treatment. Thus, once resistance to a particular drug arises, the patient may always carry that resistant viral strain (Siliciano et al., 2004, J. Antimicrobial Chemo. 54:6-9).
Commonly used anti-HIV therapies generally target two viral enzymes that HIV needs to reproduce: reverse transcriptase and protease. A third therapeutic approach is to target the binding of the virus to the cell, thereby affecting the viral entry phase of the HIV replication cycle. An example of the latter therapy is enfurvirtide (Fuzeon®), a linear 36 amino acid peptide that binds to the heptad repeat in the gp41 viral envelope glycoprotein on CD4+ cells. However, because of the problems associated with drug resistance and virus latency, there is a need for new therapeutic approaches based on anti-viral agents that are structurally and/or mechanistically different from the currently approved compounds. One such approach is represented by the anti-viral properties of phorbol compounds prostratin (see, e.g., U.S. Pat. No. 5,599,839) and ingenol (see, e.g., Fujiwara et al., 1996, Antimicrob Agents Chemother. 40(1):271-3).
Prostratin, 12-deoxyphorbol 13-acetate, was first isolated from Pimelea prostrata, a New Zealand plant toxic to livestock (Zayed S., 1977, Experentia 33(12):1554-5). Prostratin was subsequently isolated from Homalanthus nutans, a medicinal plant used by traditional Samoan healers, and shown to inhibit HIV-induced cell killing and viral replication in a variety of cell systems (Cox et al, 1993, J. Ethnopharmacol. 38(2-3):181-8; Cox, P.A., 1994, Ciba Found. Symp. 185:25-36). The potency and degree of cytoprotection is dependent on both viral strain and host cell type (Gustafson et al, 1992, J. Med. Chem. 35(11):1978-86).
Similarly, ingenol compounds were used in traditional medicine for the treatment of skin conditions (e.g., warts, corns, etc.), cancer, and asthma (Green et al., 1988, Australian J Dermatol 29:127-30 and Weedon et al., 1976, Med J of Australia 1:928). Ingenol-3,5,20-triacetate has also been show to have anti-viral properties (Fujiwara et al., supra).
Interestingly, studies suggest that prostratin displays a unique dual effect on HIV biology: inhibiting HIV replication while activating dormant or “latent” HIV that hides in human cells (Kulkosky et al., 2001, Blood. 98(10):3006-15). Prostratin efficiently reactivates HIV expression from latently infected cells generated in a SCID-hu mouse. Reactivation is associated with induction of viral transcription from the HIV long terminal repeat (LTR) and is thought to involve prostratin's property of activating specific protein kinase C (PKC) isozymes. Prostratin also appears to inhibit the entry step of the HIV replication cycle by interacting with a cellular target necessary for viral entry and/or by downregulating HIV co-receptors CCR5 and CXR5 (Witvrouw et al, 2003, Antivir Chem. Chemother. 14(6):321-8). Prostratin's unique mechanism of action is indicated by its effectiveness against different strains of HIV-1, such as HIV subtypes B and D, clinical HIV isolate (L1), HIV-2 (ROD and EHO), and SIV (MA C 251) and effectiveness against HIV strains resistant to polyanionic binding inhibitor dextran sulfate, the fusion inhibitor enfuvirtride, nucleoside reverse transcriptase inhibitors (NRTs), and protease inhibitor (PIs) (Witvrouw et al., supra). Likewise, ingenols with antiviral activity appear to affect the viral absorption process rather than the viral replication machinery (Fugiwara et al., supra). Studies further suggest that ingenol compounds may also cause reactivation of latent viruses, similar to the effects seen with prostratin (Fujiwara et al., 1998, Arch Virol. 143(10):2003-10)
Although prostratin and related phorbol compounds present an attractive therapeutic strategy for HIV in view of their reactivation and antiviral properties, as well as treatments for other diseases affected through phorbol mediated signal transduction pathways, the therapeutic effectiveness of these phorbol compounds is limited by their low solubility, low oral bioavailability, and low therapeutic index.